Current trends in conservative and minimally invasive dentistry (MID) emphasize the reversal and repair of the active caries process as a first step to restoring the diseased tissue. Enamel remineralization is an accepted phenomenon with established mechanisms, but dentin remineralization strategies are at an early stage of development. Results from the prior period have shown substantial recovery of the hydrated carious tissues mechanical properties, which we have termed functional remineralization. If functional remineralization can be clinically achieved, it would become a key strategy in MID with the eventual outcome of improved oral health care and lower costs. In support of this goal, , new knowledge on basic biomineralization mechanisms has emerged, and has inspired new approaches that achieve appropriate remineralization within collagen fibrils (intrafibrillar) and between the fibrils (extrafibrillar) to improve the functional remineralization of carious dentin structures. We propose to continue the UCSF-Univ. of Florida collaboration established in the prior period, which will include: Aim 1 a) enhancing the polymer-induced liquid-precursor (PILP) process that has successfully mineralized a variety of collagen matrices, and shown significant functional remineralization of artificial caries lesions. b) extending PILP by potential synergisti approaches including other polyanionic polymers and constant composition methods. c) studying the structure of carious dentin zones to determine their role in limitations of the remineralization process. d) defining microstructural variations in normal and functionally remineralized dentin. Aim 2 evaluates dentin collagen mineralization in model systems including mouse models that lack critical non-collagenous proteins to gain insight into mineralization mechanisms of the collagen scaffold. Aim 3 a) applies the improvements from Aims 1 and insights from Aim 2 to applications in two in vitro models of natural human caries that progressively move towards clinical application. Aim 4 establishes that functional remineralization as indicated by AFM-based nanoindentation testing of hydrated tissue also reflects properties at clinically relevant sizes by use of 4-point bending tests combined with Micro X- ray Computed Tomography. To carry out this work we have established a talented team of SF Bay area investigators from UCSF, LBNL and SSRL, as well as our Florida collaborators who developed the PILP process. The proposed studies will build on our progress and translate the emerging understanding of mechanisms in biomineralization so that we can optimize remineralization kinetics and restoration of dentin caries and move toward a clinically relevant delivery system. Establishing methods to functionally remineralize dentin caries thereby restoring the mechanical properties of the hydrated tissue will minimize conventional restorative treatment and maximize conservation of tooth structure.